Anal. Chem. 1999, 71, 3603-3609
Ion Chromatography Coupled with Mass Spectrometry for the Determination of Ionic Compounds in Agricultural Chemicals Sheher Bano Mohsin
Bayer Corporation, 8400 Hawthorn Road, Box 4913, Kansas City, Missouri 64120-2301
An IC/MS method was developed using thermospray and electrospray interfaces to obtain the mass spectra of ionic compounds in agricultural chemicals. This is an easy, rugged, and sensitive method and offers several advantages over the conventional HPLC/MS with ion-pairing reagents for the analysis of highly polar compounds. Ion exchange chromatography was used to separate the ionic compounds from other components in the sample. Sodium hydroxide mixed with methanol and water was used as the eluent. To make the eluent containing sodium hydroxide compatible with MS, an on-line solid-phase chemical suppressor was used, which removed sodium ions from the mobile solvent before it entered the mass spectrometer. The suppressor is easy to use, is rugged, and can withstand the high back-pressure generated by the thermospray LC/MS interface. It also works well with the electrospray interface.
An IC/MS method was developed using the thermospray and the electrospray LC/MS interfaces to obtain the mass spectra of ionic compounds. The biggest challenge in obtaining the mass spectra of ionic compounds using HPLC/MS is the choice of ion-pair reagents which are compatible with the mass spectrometer.1 The ion-pair reagents have to be volatile. This limits the choice of reagents which can be used. Very often, separation has to be compromised for mass spectrometer compatibility, resulting in poor peak shape and resolution. Ion exchange chromatography is a good technique for the separation of ionic compounds. However, the eluents most often used in ion exchange chromatography are not compatible with the mass spectrometer. Mobile solvents containing high salt concentrations are used in IC, making it difficult to interface the two techniques. MS does not tolerate high salt concentrations. To make the IC eluent compatible for MS, a desalting device, also called a suppressor can be used, which removes salts from the mobile solvent before it enters the mass spectrometer. Some work has been done in this area, mostly for the analysis of carbohydrates.2 Membrane suppressors have been used in the (1) Voyksner, R. D.; Haney, C. A. Anal. Chem. 1985, 57, 991. (2) Niessen, W. M. A.; Van der Hoeven, R. A. M.; Van der Greef, J.; Schols, H. A.; Varagen, A. G. J. Rapid Commun. Mass Spectrom. 1992, 6, 197. 10.1021/ac981451t CCC: $18.00 Published on Web 07/03/1999
© 1999 American Chemical Society
Table 1. Ion Chromatography Conditions for IC/MS analytical column eluent, 70% E1/30% E2 eluent flow rate sample loop volume
IonPac AS11, 250 mm × 4 mm, with IonPac AG11 guard column E1: 10 mM NaOH E2: methanol/water (80:20) 1-1.5 mL/min 50 µL
past to clean up the salts in the IC eluent before it enters the mass spectrometer.3,4 The disadvantage of using membrane suppressors is the high back pressure generated by the thermospray probe or particle beam interface, which can rupture the membrane or cause it to leak.5,6 This makes it difficult to use this technique for practical purposes. This paper reports the successful use of a solid-phase chemical suppressor for routine IC/MS work. The suppressor is rugged and can withstand high back-pressures generated by the thermospray interface. It also works well with the electrospray interface. EXPERIMENTAL SECTION The Mass Spectrometer. The mass spectrometer used is a VG (Micromass) analytical model Autospec-Q, with a VG OPUS data system and a VG SIOS interface between the VAX-based data system and the mass spectrometer. The mass spectrometer has a trisector, double-focusing geometry of the type ESA-magnetESA, and a mass range at full accelerating voltage (8 kV) of 4500 Da. An RF-only gas cell followed by a quadrupole analyzer is situated after the first detector. The RF-only gas cell can be used to produce fragments of the sample ions, which can then be analyzed by the quadrupole analyzer. The entire unit is of EBEQQ geometry. The thermospray and the electrospray interfaces were obtained from Micromass. (3) Simpson, R. C.; Fenselau, C. C.; Hardy, M. R.; Townsend, R. R.; Lee, Y. C.; Cotter, R. B. Anal. Chem. 1990, 62, 248. (4) Conboy, J. J.; Henion, J. D.; Martin, M. W.; Zweigenbaum, J. A. Anal. Chem. 1990, 62, 800. (5) Torto, N.; Hofte, A.; Van der Hoeven, R.; Tjaden, U.; Gorton, L.; MarkoVarga, G.; Bruggink, C.; Van der Greef, J. J. Mass Spectrom. 1998, 33, 334. (6) Niessen, W. M. A.; Van der Hoeven, R. A. M.; Van der Greef, J.; Schols, H. A.; Varagen, A. G. J. Rapid Commun. Mass Spectrom. 1992, 6, 197. (7) Cairns, T.; Siegmund, E. G.; Stamp, J. J. Mass Spectrom. Rev. 1989, 8, 127.
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Figure 1. Schematic diagram of the IC/MS system. Table 2. Mass Spectrometer Conditions for Thermospray LC/MS interface source temperature
thermospray 200°C
probe tip temperature source pressure
quadrupole resolution collision gas
1 air at 5 × 10-5 mbar
Quadrupole Parameters collision energy 76 eV quadrupole scan range 150-20
function
mass (amu)
1 2
144.0 130.0
210°C 4 × 10-4 mbar
Daughter Quadrupole Experimental Parameters scan range mass (amu) resolution time (min) mode 150-20 150-20
1000 1000
The IC System. IC separations were performed on a Dionex model 4000i ion chromatograph, equipped with an anion exchange IonPac AS11 analytical column, an IonPac AG11 guard column, and a conductivity detector. The AS11 anion exchange column is specially designed for using hydroxide eluent systems. It is stable between pH 0 and pH 14 and is compatible with eluents containing 0-100% organic solvents. The IC conditions used are shown in Table 1. The Autosuppressor. The Alltech 1000HP electrochemically regenerated ion suppressor (ERIS) is a solid-phase chemical suppressor. It was installed between the analytical column and the detector. It consists of two cells packed with a strong cation exchanger in the hydrogen form. The sodium ions from the mobile solvent are exchanged with hydrogen ions in the cell to form water which goes to the detector. The analytes are converted to their acid forms and can be detected using a conductivity detector or a mass spectrometer. A 10-port valve is used to switch mobile solvent between the two cells. While one cell is used to suppress the mobile phase, the other cell is electrochemically regenerated to produce the hydrogen form of the resin. Experimental Setup for Thermospray. The IC column was connected to the autosuppressor unit. The effluent from the autosuppressor was mixed with 80:20 methanol/water containing 0.05 M ammonium acetate before it went to the thermospray unit. An HPLC pump was used to deliver the methanol/water containing ammonium acetate. Ammonium acetate is needed for thermospray operation. A T-connector was used to connect the two mobile solvent lines to the thermospray unit. The IC system was placed on a cart so that it could be moved close to the mass 3604 Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
1.5 1.5
TS+ TS+
accelerating voltage MS resolution
8 kV 1000
start time (min)
end time (min)
0.01 4.06
4.00 10.00
spectrometer when needed. A schematic diagram of the IC/MS system is shown in Figure 1. The mass spectrometer was tuned to a resolution of 1000. The instrument was operated in the MS/ MS mode, and daughter ions resulting from the M + 18 ion of the analyte were recorded. Table 2 shows the MS conditions used for thermospray. Experimental Setup for Electrospray. The flow rates generally used for electrospray are in the 10-50 µL/min range. Because of this requirement, effluent from the suppressor was split 99:1 and mixed with 90:10 acetonitrile/water containing 0.5% ammonium hydroxide before it went to the electrospray probe unit. The electrospray was operated in the negative-ion mode. A syringe pump was used to deliver acetonitrile/water containing ammonium hydroxide. Ammonium hydroxide increases the pH of the mobile solvent and assists in negative-ion formation. Calibration compounds were also introduced into the mass spectrometer using the syringe pump. The region between the sampling cone and the skimmer is an intermediate-pressure region in the electrospray source. Sample ions can gain sufficient energy to undergo collision-induced dissociation (CID) reactions with neutral molecules in this region, resulting in fragment ions. The voltage difference between the sampling cone and the skimmer was increased to assist in fragmentation of the [M - H]- ions. Table 3 shows the MS conditions used for electrospray. RESULTS AND DISCUSSION Figure 2 shows the structures of the compounds identified in the sample. Figures 3-8 are the results obtained with the thermospray interface. Figure 3 shows the chromatograms
Table 3. Typical Mass Spectrometer Conditions for Electrospray ion mode needle voltage sampling cone voltage skimmer lens voltage ring electrode voltage bath heat scan range scan time accelerating voltage resolution
negative -8000 V -4050 V -4000 V -3980 V 80 °C 50-600 amu 1.5 s/decade 4000 V 1000
Figure 2. Structures of methyl phosphate, methyl sulfate, and dimethyl phosphate.
obtained from a standard and a sample injection using the conductivity detector. The standard was a mixture of approximately 10 ppm of dimethyl phosphate and 10 ppm of methyl phosphate. Both of these components are shown in the standard chromatogram in Figure 3. Dimethyl phosphate, which has a single negative charge, elutes during the first 3 min, followed by the doubly charged methyl phosphate ion. Methyl phosphate elutes after the first 4 min under these conditions. The sample chromatogram of an organophosphate insecticide obtained by using the conductivity detector is shown in the bottom panel of Figure 3. A number of components elute close to dimethyl phosphate during the first 3 min. Methyl phosphate elutes after 4 min and therefore has no interference from other peaks. Figure 4 shows the chromatograms of the standard resulting from using the mass spectrometer for detection. The experimental setup for the MS/MS experiment was as follows: During the first 4 min of the experiment, m/z 144 (the M + 18 ion of dimethyl phosphate) was transmitted by the ESAmagnet-ESA to the RF-only collision cell, and the ions generated from the dissociation of the parent ion were recorded by the quadrupole analyzer. The top trace in Figure 4 shows the total ion chromatogram resulting from the fragmentation of the m/z 144 ion for the first 4 min. The top chromatogram therefore has data recorded only for 4 min. The peak at ∼2 min is due to dimethyl phosphate in the standard. After the first 4 min, the m/z 130 ion (the M + 18 ion of methyl phosphate) was transmitted through the ESA-magnet-ESA to the collision cell, and its daughter ions were recorded by the quadrupole analyzer. The bottom chromatogram shows data acquired from 4 to 8 min. It therefore shows no data for the first 4 min. The bottom chromatogram in Figure 4 is due to the fragmentation of the m/z 130 ion. The peak at ∼5.5 min is due to methyl phosphate in the standard. Figure 5 shows the MS/MS spectrum of dimethyl phosphate in the standard. The major peaks in the MS/MS spectrum of dimethyl phosphate are the m/z 144/145 and m/z 127 ions. The peak at m/z 127 is due to the addition of a proton to the molecular
Figure 3. Chromatograms of the standard (top) and sample (bottom) using conductivity detection.
ion (M + H). A low-intensity peak is also seen at m/z 95 (PO4 ion). Figure 6 shows the MS/MS spectrum of methyl phosphate in the standard. Major peaks are seen at m/z 130 and m/z 113. The peak at m/z 113 results from the M + H ion. It also shows a lowintensity peak at m/z 95 due to the phosphate ion. Figure 7 shows MS/MS chromatograms resulting from injecting a sample of 1% technical organophosphate insecticide. The top chromatogram, due to the fragmentation of the m/z 144 ion (the M + 18 ion of dimethyl phosphate) shows a lot of interference from other impurity peaks, consistent with the chromatogram recorded with conductivity detection (Figure 3). The bottom chromatogram shows a single peak due to methyl phosphate. The retention time of ∼5.5 min and the MS/MS spectrum of this peak are very similar to those of methyl phosphate in the standard. Figure 8 shows the MS/MS spectrum of methyl phosphate in the sample. The MS/MS spectrum of dimethyl phosphate in the sample could not be obtained due to interference problems. A better separation is being developed. Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
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Figure 4. Total ion chromatograms of dimethyl phosphate (top) and methyl phosphate (bottom) in the standard using the mass spectrometer for detection.
Figure 5. MS/MS of dimethyl phosphate in the standard. 3606 Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
Figure 6. MS/MS of methyl phosphate in the standard.
Figure 7. Total ion chromatograms showing the presence of methyl phosphate in the sample using the mass spectrometer for detection.
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Figure 8. MS/MS of methyl phosphate in the sample.
Figure 9. Ion chromatogram resulting from the m/z 111 ion recorded with electrospray.
Figures 9-11 show the results obtained using the electrospray interface. Figure 9 shows the chromatogram due to the m/z 111 ion. Peaks due to methyl sulfate and methyl phosphate in an organophosphate sample are seen in the chromatogram. The [M - H]- ions resulting from methyl sulfate and methyl phosphate have the same m/z of 111, yet their mass spectra are different. The mass spectrum of methyl phosphate, shown in Figure 10, has a fragmentation pattern different from that of the mass spectrum of methyl sulfate shown in Figure 11. The mass spectrum of methyl sulfate clearly shows the typical isotopic cluster resulting from the presence of sulfur in this compound. 3608 Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
Figure 10. Mass spectrum of methyl phosphate recorded with electrospray.
The isotopic cluster around m/z 111 for methyl phosphate is consistent with the lack of sulfur in this compound. CONCLUSIONS IC/MS was successfully used for the analysis of ionic compounds in agricultural chemicals. The solid-phase suppressor makes it possible to interface the ion chromatograph to the mass spectrometer. The thermospray mass spectra of methyl phosphate and dimethyl phosphate are very simple with exclusively an M + 18 peak. The M + 18 ion was selectively transmitted to the collision cell and fragmented, and its daughter ion mass spectrum was recorded using the quadrupole analyzer.
[M - H]- peak at 111 m/z. The [M - H]- peak was fragmented in the intermediate-pressure region of the electrospray source to give fragment ions. Methyl sulfate and methyl phosphate can be easily differentiated from their mass spectra. The IC/MS technique using the solid-phase suppressor for online desalting of the mobile solvent is simple and rugged and can be used routinely for the mass spectrometric analysis of ionic compounds. ACKNOWLEDGMENT
Figure 11. Mass spectrum of methyl sulfate recorded with electrospray.
Mass spectra of methyl phosphate and methyl sulfate in an organophosphate matrix were also recorded using the electrospray interface. Methyl phosphate and methyl sulfate give an intense
I wish to thank Mr. Steve Slahck, Mr. Fred Smead, and Mr. Carl Gregg for their encouragement and support during this project. I also wish to thank Dr. Sarah Leibowitz for her help in preparing the manuscript.
Received for review December 31, 1998. Accepted May 25, 1999. AC981451T
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